Background and Objective: E-learning has become an increasingly popular learning approach in higher educational institutions due to the rapid growth of technologies. We need valid and reliable tools to assess the acceptance of this type of training. This study aimed to validate the e-learning acceptance scale.Materials and Methods: This psychometric study was conducted on 223 samples of virtual education students from the Medical Universities in Tehran. Initial instrument was 49 questions from previous studies that were translated into Persian after obtaining permission from the original author. Instrument validation procedures included impact item method, content validity index (CVI) and content validity ratio (CVR) of the judgment of 10 experts and exploratory factor analysis and internal reliability of the scale was determined by Cronbach's alpha and with the test-retest method the external consistency of a test was assessed.Results: Of the original 49 items, all of them based on the impact item index, CVR, and CVI were considered as valid; based on exploratory factors analysis, 41 items were retained. And considering a minimum Eigen value of 1 for each factor, the five factors were extracted. This instrument was able to predict 52% of the properties as an acceptance e-learning variance.Conclusion: The results of this study indicated the strength of the factor structure and reliability of an instrument for measuring student acceptance e-learning scale.